Size Exclusion Chromatography Chromatograms Research Articles

Electrochemical oxidation treatment of reverse osmosis concentrated landfill leachate: Effect of operation parameters and evolution of dissolved organic matter

Electrochemical oxidation (EO) is a viable option for the advanced treatment of reverse osmosis concentrated landfill leachate (ROCLL). This work characterized dissolved organic matter (DOM) in ROCLL with size exclusion chromatography (SEC) and then investigated the impact of operating parameters on the removal of DOM fractions with response surface methodology (RSM). The SEC chromatograms revealed that the DOM fractions in ROCLL comprised of biopolymers (BP, > 18 kDa), humic substances (HS, 0.45–18 kDa) and low molecular weight substances (LMWS, < 0.45 kDa). For the EO configuration with different anodes, the BDD film anode showed the highest removal efficiency for DOM in terms of COD, DOC, UVA254 and fluorescence indices, followed by Ti, graphite, Ti/RuO2-IrO2 and Ti/SnO2-Sb2O5 anodes. The RSM experiments suggested that pH was the most critical factor to enhance the removal of all DOM fractions, and acidification pretreatment could remove approximately 50% BP and 30% HS fractions due to the pH-induced precipitation. The DOC removal kinetics of DOM fractions could be described by pseudo-zero kinetic model with rates in the order of HS > LMWS > BP, which were controlled by charge transfer process. Differently, the elimination of chromophores and fluorophores followed pseudo-first kinetics, which was limited by mass transport process. Via nonnegativity matrix factorization (NMF) deconvolution of the SEC-OCD chromatograms, BP-C1, HS-C2 and LMWS-C1 subfractions required to be removed through further treatment after EO process. This work demonstrates the potential for SEC analysis coupled with mathematical approaches to give informative insights into the evolution of DOM and provide optimization strategies for the EO treatment of complex ROCLL.

Biochemical and biophysical characterization of the RAS family small GTPase protein DiRAS3

DiRAS3, also called ARHI, is a RAS (sub)family small GTPase protein that shares 50–60% sequence identity with H-, K-, and N-RAS, with substitutions in key conserved G-box motifs and a unique 34 amino acid extension at its N-terminus. Unlike the RAS proto-oncogenes, DiRAS3 exhibits tumor suppressor properties. DiRAS3 function has been studied through genetics and cell biology, but there has been a lack of understanding of the biochemical and biophysical properties of the protein, likely due to its instability and poor solubility. To overcome this solubility issue, we engineered a DiRAS3 variant (C75S/C80S), which significantly improved soluble protein expression in E. coli. Recombinant DiRAS3 was purified by Ni-NTA and size exclusion chromatography (SEC). Concentration dependence of the SEC chromatogram indicated that DiRAS3 exists in monomer-dimer equilibrium. We then produced truncations of the N-terminal (ΔN) and both (ΔNC) extensions to the GTPase domain. Unlike full-length DiRAS3, the SEC profiles showed that ΔNC is monomeric while ΔN was monomeric with aggregation, suggesting that the N and/or C-terminal tail(s) contribute to dimerization and aggregation. The 1H–15N HSQC NMR spectrum of ΔNC construct displayed well-dispersed peaks similar to spectra of other GTPase domains, which enabled us to demonstrate that DiRAS3 has a GTPase domain that can bind GDP and GTP. Taken together, we conclude that, despite the substitutions in the G-box motifs, DiRAS3 can switch between nucleotide-bound states and that the N- and C-terminal extensions interact transiently with the GTPase domain in intra- and inter-molecular fashions, mediating weak multimerization of this unique small GTPase.

Iron-induced association between selenium and humic substances in groundwater from deep sedimentary formations

ABSTRACT To better understand the impact of natural organic matters (NOM) on facilitating 79Se migration in underground environments, the association mechanism between Se and humic substance (HS) in groundwater was investigated through incubation experiments using groundwater samples under abiotic and biotic conditions and speciation analyses using a DAX-8 resin isolation technique and a size exclusion chromatography (SEC) coupled online to ultraviolet–visible (UV–Vis) and inductively coupled plasma mass spectrometry (ICP-MS) detection (SEC-UV-ICP-MS). Selenite added into the groundwater was reduced to elemental Se only when microbial activity was stimulated by a nutrient. The DAX-8 resin isolation technique showed that Se associated with HS in both the oxidizing and reducing conditions, but the proportion of the associated Se to the total dissolved Se was the highest under the reducing condition. The SEC-UV-ICP-MS analyses of Se, Fe, and HS extracted from the DAX-8 resin showed that there is a fraction of Se that simultaneously elutes with Fe and HS. Logarithmic peak areas of Se and Fe associated with HS under the moderately oxidizing conditions were linearly correlated regardless of the presence of microorganisms and nutrient ( r = 0.947, p ≤ 0.01 with the addition of Se; r = 0.995, p < 0.01 without the addition of Se). The SEC chromatogram of Se associated with HS under the reducing condition was well-matched with those under the moderately oxidizing conditions. Therefore, these suggest that Fe-induced association between Se and HS can preferentially occur in the groundwater conditions.

Multivariate correlation of infrared fingerprints and molecular weight distributions with bioactivity of poultry by-product protein hydrolysates

• Processing conditions influence the bioactive properties of protein hydrolysates. • FTIR and SEC fingerprints are correlated with bioactivities of protein hydrolysates. • PLSR models predicted ACE-1 inhibition and DPPH radical scavenging activity. • FTIR-based models were observed to be more robust than SEC-based models. Characterization of protein hydrolysates is a vital step in developing peptide-based bioactive ingredients. Multivariate correlation of chemical fingerprints and bioactivity of poultry by-product protein hydrolysates is explored as a potential analytical strategy for characterization and quality control. Chemical fingerprints of sixty hydrolysates were acquired using Fourier-transform infrared spectroscopy (FTIR) and size exclusion chromatography (SEC). Bioactivities (2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and angiotensin-1-converting enzyme (ACE-1) inhibition) were measured in vitro . Partial least squares regression models based on FTIR fingerprints or SEC chromatograms showed a better prediction performance for ACE-1 inhibition (coefficients of determination (R 2 ) = 0.91, root mean square error of prediction (RMSECV) = 2.8; R 2 = 0.85, RMSECV = 3.5, respectively) than for DPPH radical scavenging (R 2 = 0.74, RMSECV = 0.3; R 2 = 0.75, RMSECV = 0.3, respectively). Such models are promising tools for rapid prediction of bioactivities and as a quality control technology in production of bioactive peptides.

Characterizing Molecular Weight Distribution and Optical Properties of Dissolved Organic Matter and Unraveling the Origins of Anthropogenic Fluorophores in Yangtze River and Its Tributaries

In this study, size exclusion chromatography (SEC) coupled with multiple detectors was used to characterize the composition, molecular weight (MW), and optical properties of dissolved organic matter (DOM) in the Yangtze River main stream and its 88 tributaries. The SEC chromatograms revealed that DOM in the investigated water samples mainly comprised nonhumic proteinaceous biopolymers with apparent MW > 20 kDa and humic substances with apparent MW ranging from ∼0.5 to ∼7 kDa. The SEC fluorescence chromatograms further unraveled that there was a kind of characteristic fulvic-acid-like fluorophore in downstream tributaries, which showed triple-excitation peaks at ∼250, 310, and 365 nm with an emission peak at ∼460 nm (designated as HS-Em460-Ex3 hereinafter). Via field investigation and parallel factor analysis, this characteristic HS-Em460-Ex3 fluorophore was identified to originate from the discharge of residual naphthol and naphthylamine moieties in biologically treated textile wastewater. The SEC fluorescence fingerprints also demonstrated the occurrence of the characteristic HS-Em460-Ex3 fluorophore in the Yangtze River main stream from Changzhou to Nantong, suggesting that the discharge of textile wastewater might have altered the optical properties of riverine DOM and advanced treatment processes should be applied to eliminate residual aromatic fluorophores in textile wastewater.

Resolving humic and fulvic acids in binary systems influenced by adsorptive fractionation to Fe-(hydr)oxide with focus on UV–Vis analysis

Humic acid (HA) and fulvic acid (FA) are two operationally defined classes of natural organic matter. In the environment, both materials are present simultaneously and bind in a competitive manner to Fe-(hydr)oxides and other minerals, but their quantification in mixtures is a challenge. In this study, an UV–Vis method was developed to quantify concentrations of HA and FA without and after adsorptive fractionation by an iron oxide (goethite, α-FeOOH). In addition, the performance of the UV–Vis method was compared to that of acid precipitation and size exclusion chromatography (SEC). Among the three methodologies (UV–Vis, acid precipitation, SEC), the UV–Vis method is the most successful in quantifying the ratio of HA to FA subject to fractionation. The UV–Vis method is based on distinct differences in the UV–Vis spectra of HA and FA, including fingerprints in both the spectra shape and intensity. Adsorption to goethite decreased the specific light absorbance of HA and FA, but the changes in spectral shape were not significant enough to cover their differences. The acid precipitation method can also quantify the HA to FA ratio. But to minimize the influence of incomplete HA precipitation or co-precipitation of FA, the concentration of both HA and FA needs to be at least ~20 mgC L−1. The SEC method is not suitable to measure HA and FA after adsorption, because preferential adsorption significantly affects the shape of SEC chromatograms.

Enzymatic crosslinking of silk sericin through combined use of TGase and the custom peptide

Silk sericin (SS) is derived from natural Bombyx mori silk fibers, and it has the unique biological properties like good biocompatibility and low immunogenicity. However, pure SS material exhibits poor mechanical properties and high water solubility, which greatly reduces the formability of SS material and limits its wide range of applications. In this study, a transglutaminase (TGase) was combinedly used with two custom peptides of GQGEGQG (p-Q) and KKKK (p-K), containing exogenous glutamine and lysine residues, respectively, for facilitating the enzymatic crosslinking of sericin proteins. The efficacy of enzymatic incubation was assessed by the means of SDS-PAGE, size exclusion chromatography chromatogram, and determination of the quantitative changes in amino groups. Properties of the obtained SS membranes based on different treatments were also examined. The results revealed that incubating of SS with TGase and glutamine-containing peptide led to a noticeable increase in the molecular weight compared with others. TGase-mediated treatments had detectable impact on the conformational structures of SS, the obtained formability and mechanical property for the TGase/p-Q treated SS membrane were evidently improved compared to others. The suggestive alternative, based on the combination use of TGase and p-Q has the potential applications for preparation of the sericin-based biomaterials.

Off-line comprehensive size exclusion chromatography × reversed-phase liquid chromatography coupled to high resolution mass spectrometry for the analysis of lignocellulosic biomass products

Biochemical and thermochemical processes are two pathways to convert lignocellulosic biomass into fuels and chemicals. Both conversion types produce aqueous complex samples containing many oxygenated chemical functions over a wide range of masses. Nowadays, composition of these biomass products is still largely unknown, especially their nonvolatile part (300–1000 Da) mostly made of carbohydrates and their derivatives. In the present study, size exclusion chromatography (SEC) was investigated and applied on water soluble phase of a fast pyrolysis bio-oil (thermochemical conversion) and on aqueous phase of pretreated wheat straw (biochemical conversion). An optimization of mobile phase composition using model molecules was necessary to limit non-steric interactions and elute all chemical families. At the end, separation of carbohydrates, heterosides and aromatic species was performed. The chemical organization of SEC chromatograms was confirmed by coupling SEC with a Fourier transform-ion cyclotron resonance mass spectrometer (FT-ICR MS) using electrospray ionization (ESI) in the negative mode. On-line SEC-UV/FT-ICR MS hyphenation was a powerful tool to provide exact mass distribution of samples and get molecular formulae classed by chemical family. To go further, the complementarity of SEC with reversed-phase liquid chromatography (RPLC) was established with an off-line comprehensive 2D-LC analysis of the two samples. First, 140 fractions were collected physically from SEC separation for each sample, then each fraction was analyzed by RPLC hyphenated to an Ion Trap - Time of Flight mass spectrometer (SEC × RPLC-UV/IT-TOF MS) using ESI in both positive and negative modes. This comprehensive approach combining 2D-LC and high resolution mass spectrometry nearly doubled the number of peaks detected in comparison with 1D RPLC analysis and thus offered well resolved 2D contour plots, considered as relevant analytical fingerprints of the aqueous phase of biomass samples.

A new molecular weight (MW) descriptor of dissolved organic matter to represent the MW-dependent distribution of aromatic condensation: Insights from biodegradation and pyrene binding experiments

In this study, we utilized a size exclusion chromatography (SEC) system that was equipped with a fluorescence emission scanning mode to explore the heterogeneous distribution of the humification index (HIX) values within bulk dissolved organic matter (DOM). The HIX-based SEC chromatograms showed that the molecular weight (MW)-dependent distribution of aromatic condensation was heavily affected by the DOM sources and the progress of biodegradation. The HIX heterogeneity across different MW was more pronounced for terrestrial versus aquatic DOM sources. Microbial incubation of leaf litter DOM led to the initial enhancement of the HIX at a relatively low MW, followed by a gradual increase at larger MW values. The dynamic changes of the HIX can be attributed to (1) the preferential removal of non-aromatic or less-aromatic molecules by microorganisms, (2) the production of microbial metabolites, (3) microbial humification, and (4) self-assemblage of humic-like molecules. From the SEC chromatograms, the HIX-based average molecular weight (or MWHIX) was proposed as a unifying surrogate to represent an MW that was highly associated with aromatic condensation. The MWHIX discriminated four different DOM sources and described well the biodegradation-induced changes. The MWHIX also presented a good positive correlation with pyrene organic carbon-normalized binding coefficients (Koc). The prediction capability of the MWHIX for pyrene Koc was higher than those based on the single descriptors of bulk DOM, such as HIX and MW, which revealed its superior linkage with the DOM reactivity related to both MW and HIX.

Solubilization of cashew gum from Anacardium occidentale in aqueous medium

Although cashew gum (CG) is known to be soluble in water, the solubilized CG does not exist as individual molecules in its solution. Instead, CG molecules form aggregates resulting in a turbid solution. For better solubilization of CG in water, two types of approaches are attempted: thermal degradation and changing the pH of solution medium. The thermal degradation at high temperatures does not follow the same pattern as a similar polysaccharide, starch: instead of being thermally degraded to smaller molecules that are readily soluble in water, the aggregated CG molecules were not fully disintegrated to individual molecules even after the heating to 190 °C. Size Exclusion Chromatography (SEC) shows that CG exists as entangled aggregates in aqueous solution at room temperature, and these aggregates disentangle in neutral or alkaline medium. Therefore, raising the pH of solution turns out to be a more efficient way to dissolve CG in water than thermal degradation. CG aggregates disintegrate only when the pH of its solution is adjusted to higher than 6.2. The peak MW of CG is measured to be 28 kDa. The lowest temperature that will initiate the thermal degradation and the temperature that induces full degradation of CG are estimated from the SEC chromatogram of heat-treated CG’s.

Preliminary Characterization and Bioactivities of Some Impatiens L. Water-Soluble Polysaccharides.

Preliminary characterization and bioactivity of water-soluble polysaccharides from four Impatiens species—I. glandulifera Royle, I. parviflora DC., I. balsamina L., and I. noli-tangere L.—were investigated. The yields of polysaccharides range widely from 1.97% for I. parviflora roots to 18.63% for I. balsamina aerial parts. SEC (Size exclusion chromatography) chromatograms show that all samples contained a low molecular weight part that consisted of components of similar molecular weight. The aerial parts and roots of I. balsamina, and I. glandulifera aerial parts had considerable amounts of high molecular weight components up to 2.3 MDa. The sugar composition analysis revealed that Impatiens polysaccharides consisted primarily of galactose, arabinose, rhamnose, mannose, xylose, and glucose. All polysaccharide fractions, except for I. parviflora roots, also contain galacturonic acid. Moreover, in vitro bioactivity of obtained polysaccharides were evaluated. The antioxidant activity was evaluated on the basis of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azino-bis-(3-ethyl-benzthia-6-sulfonic acid) (ABTS) radical scavenging assays. The highest antioxidant activity was obtained for I. balsamina aerial parts and I. parviflora roots. Among the tested fractions, only the polysaccharides from I. glandulifera aerial parts were able to significantly decrease the production of IL-8 by 32.7 ± 10.5%. The results suggest that Impatiens species can be considered as a new source of antioxidants.

The Use of Size Exclusion Chromatography to Monitor Protein Self-Assembly

High resolution size exclusion chromatography (SEC) coupled with static light scattering (SLS) analyses were conducted to study the effect of the mobile phase ionic strength and protein concentration on the output of SEC experiments. The results highlight the effect of small changes in the mobile phase composition on the estimation of molar masses estimated from retention time-based calibration curve compared with those obtained from SLS analysis. By comparing the SLS data with the SEC chromatograms, we show that SEC can provide helpful information on the protein aggregation state as macromolecules approach known precipitation points in their phase diagrams. This suggests the potential use of SEC as an easily accessible lab-based scanning methodology to monitor protein self-assembly prior to nucleation and crystallization. Implications for the use of SEC to study protein phase diagrams are discussed.

Quality of bevacizumab (Avastin®) repacked in single-use glass vials for intravitreal administration.

Avastin® (bevacizumab) is an anti-vascular endothelial growth factor (VEGF) monoclonal antibody given as an off-label drug by intravitreal administration for treatment of ocular diseases. The drug's clinical application and its cost-benefit profile has generated demand for its division into single-use vials to meet the low volume and low-cost doses necessary for intraocular administration. However, the safety of compounding the drug in single-use vials is still under discussion. In this study, the stability and efficacy of Avastin® repacked in individual single-use glass vials and glass ampoules by external compounding pharmacies were evaluated. Polyacrylamide gel electrophoresis (PAGE), size-exclusion chromatography (SEC), dynamic light scattering (DLS), and turbidimetry were selected to detect the formation of aggregates of various sizes. Changes in bevacizumab biological efficacy were investigated by using an enzyme-linked immunosorbent assay (ELISA). Repacked and reference bevacizumab showed similar results when analyzed by PAGE. By SEC, a slight increase in high molecular weight aggregates and a reduction in bevacizumab monomers were observed in the products of the three compounding pharmacies relative to those in the reference bevacizumab. A comparison of repacked and reference SEC chromatograms showed that the mean monomer loss was ≤1% for all compounding pharmacies. Protein aggregates in the nanometer- and micrometer-size ranges were not detected by DLS and turbidimetry. In the efficacy assay, the biological function of repacked bevacizumab was preserved, with <3% loss of VEGF binding capacity relative to that of the reference. The results showed that bevacizumab remained stable after compounding in ampoules and single-use glass vials; no significant aggregation, fragmentation, or loss of biological activity was observed.

Cross-Contamination From Vial Caps for SEC Analysis Determined by MALDI-TOF Mass Spectrometry.

Cross-contamination from the polymeric material of the vial caps used in size exclusion chromatography (SEC) and high-performance liquid chromatographyautosamplers in the low molar mass (LMM) range (<2000 Da) of polymer samples was demonstrated with the help of matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) analysis. It is shown that polyethylene (PE) caps, if exposed to either short or long periods intetrahydrofuran solvent, might extract oligomers that contaminate the LMM end of the SEC chromatograms. On the other hand, polytetrafluoroethylene (PTFE)/silicone caps have shown to be inert if used properly; namely, the PTFE part in contact with the sample solution. However, care should be taken if accidentally the silicon part is exposed to the solution because silicone oligomer peaks have been detected by MALDI-TOF MS analysis that might mislead the results.

Using an FPLC to promote active learning of the principles of protein structure and purification.

The concepts of protein purification are often taught in undergraduate biology and biochemistry lectures and reinforced during laboratory exercises; however, very few reported activities allow students to directly gain experience using modern protein purification instruments, such as Fast Protein Liquid Chromatography (FPLC). This laboratory exercise uses size exclusion chromatography (SEC) and ion exchange (IEX) chromatography to separate a mixture of four different proteins. Students use an SEC chromatogram and corresponding SDS-PAGE gel to understand how protein conformations change under different conditions (i.e. native and non-native). Students explore strategies to separate co-eluting proteins by IEX chromatography. Using either cation or anion exchange, one protein is bound to the column while the other is collected in the flow-through. In this exercise, undergraduate students gain hands-on experience with experimental design, buffer and sample preparation, and implementation of instrumentation that is commonly used by experienced researchers while learning and applying the fundamental concepts of protein structure, protein purification, and SDS-PAGE. © 2016 by The International Union of Biochemistry and Molecular Biology, 45(1):60-68, 2017.

Mechanism investigation for poloxamer 188 raw material variation in cell culture.

Variability in poloxamer 188 (P188) raw material, which is routinely used in cell culture media to protect cells from hydrodynamic forces, plays an important role in the process performance. Even though tremendous efforts have been spent to understand the mechanism of poloxamer's protection, the root cause for lot-to-lot variation was not clear. A recent study reported that the low performance was not due to toxicity but inefficiency to protect cells (Peng et al., Biotechnol Prog. 2014;30:1411-1418). In this study, it was demonstrated for the first time that the addition of other surfactants even at a very low level can interfere with P188 resulting in a loss of efficiency. It was also found that the performance of P188 lots correlated well with its foam stability. Foam generated from low performing lots in baffled shaker flask lasts longer, which suggests that the components in the foam layers are different. The spiking of foam generated from a low performing lot into the media containing a high performance lot resulted in cell damage and low growth. Analytical studies using size exclusion chromatography (SEC) identified differences in high molecular weight (HMW) species present in the P188 lots. These differences are much clearer when comparing the HMW region of the SEC chromatogram of foam vs. bulk liquid samples. This study shows that low performing lots have enriched HMW species in foam samples due to high hydrophobicity, which can be potentially used as a screening assay. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:767-775, 2016.

Light-Activated, In Situ Forming Gel for Sustained Suprachoroidal Delivery of Bevacizumab

A light-activated polycaprolactone dimethacrylate (PCM) and hydroxyethyl methacrylate (HEMA) based gel network was developed to sustain the release of stable, active bevacizumab (an anti-VEGF antibody used to treat choroidal neovascularization) and used to assess sustained ex vivo delivery in rabbit eyes and in vivo delivery in rat eyes following in situ gel formation in the suprachoroidal space. PCM was synthesized from polycaprolactone diol (PCD) and evaluated using NMR spectroscopy. PCM was used to cross-link HEMA in the presence of 365 nm UV light and 2,2-dimethoxy-2-phenylacetophenone (DMPA) as a photoinitiator. Bevacizumab was entrapped in the gel using three different cross-linking durations of 3, 7, and 10 min. In vitro release of bevacizumab in PBS pH 7.4 at 37 °C during a 4 month study was quantified using a VEGF-binding based ELISA. The stability of released bevacizumab was monitored by size exclusion chromatography (SEC) and circular dichroism. Alexa Fluor 488 dye conjugated bevacizumab mixed with polymers was injected suprachoroidally in rabbit eyes to study the effect of different cross-linking durations on the spread of the dye conjugated bevacizumab. In vivo delivery was assessed in Sprague-Dawley (SD) rats by injecting Alexa Fluor 488 dye conjugated bevacizumab mixed with polymers followed by cross-linking for 10 min. Spread in the rabbit eyes and in vivo delivery in rat eyes was monitored noninvasively using a fundus camera and Fluorotron Master. The formation of PCM was confirmed by the disappearance of hydroxyl peak in NMR spectra. A cross-linking duration of 10 min resulted in a burst release of 21% of bevacizumab. Other cross-linking durations had ≥62% burst release. Bevacizumab release from 10 min cross-linked gel was sustained for ∼4 months. Release samples contained ≥96.1% of bevacizumab in the monomeric form as observed in SEC chromatograms. Circular dichroism confirmed that secondary β-sheet structure of bevacizumab was maintained after release from the gel. As the cross-linking duration was increased to 10 min, the gel/antibody was better confined at the injection site in excised rabbit eye suprachoroidal space. Delivery of Alexa Fluor 488 dye conjugated bevacizumab was sustained for at least 60 days in the suprachoroidal space of SD rats. PCM and HEMA gel sustained bevacizumab release for 4 months and maintained the stability and VEGF-binding activity of bevacizumab. Therefore, light-activated PCM and HEMA gel is suitable for in situ gel formation and sustained protein delivery in the suprachoroidal space.

Toward Understanding the Architecture (Branching and MWD) of Crosslinked Acrylic Latexes

AbstractThe architecture of crosslinked acrylic latexes of 2‐ethylhexyl acrylate, methacrylic acid, and a crosslinker agent are investigated. These polymers contain a significant amount of gel content. Branching is obtained by liquid‐ and melt‐state 13C NMR spectroscopy. For the first time, the branching densities (long and short chains) of the soluble and gel fractions are measured independently. Branching is higher for the polymers produced with the most‐efficient crosslinkers, namely those which yield a higher crosslinking density. The branching (long chains) and the molecular‐weight distribution (MWD) of the soluble fraction are characterized by size‐exclusion chromatography (SEC) coupled with triple detection. Coelution of branched chains at large elution volumes is observed in the SEC chromatograms indicating the highly branched nature of the sol fractions of the latexes.

Elution behaviour of low molecular weight compounds in size exclusion chromatography

Research Article| February 01 2012 Elution behaviour of low molecular weight compounds in size exclusion chromatography Aki Sebastian Ruhl; Aki Sebastian Ruhl 1Berlin Institute of Technology, Chair of Water Quality Control, Str. des 17. Juni 135, 10623 Berlin, Germany E-mail: aki.s.ruhl@tu-berlin.de Search for other works by this author on: This Site PubMed Google Scholar Martin Jekel Martin Jekel 1Berlin Institute of Technology, Chair of Water Quality Control, Str. des 17. Juni 135, 10623 Berlin, Germany Search for other works by this author on: This Site PubMed Google Scholar Journal of Water Supply: Research and Technology-Aqua (2012) 61 (1): 32–40. https://doi.org/10.2166/aqua.2012.138 Article history Received: May 19 2011 Accepted: November 15 2011 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Share Icon Share Facebook Twitter LinkedIn MailTo Tools Icon Tools Cite Icon Cite Permissions Search Site Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsThis Journal Search Advanced Search Citation Aki Sebastian Ruhl, Martin Jekel; Elution behaviour of low molecular weight compounds in size exclusion chromatography. Journal of Water Supply: Research and Technology-Aqua 1 February 2012; 61 (1): 32–40. doi: https://doi.org/10.2166/aqua.2012.138 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex Size exclusion chromatography (SEC) in combination with continuous organic carbon detection (OCD) is a powerful analytical technique that enables characterization of dissolved organic water constituents. Low molecular weight organic water constituents are ubiquitous but their behaviour in SEC analyses is not yet fully clarified. Therefore, we analysed a number of low molecular weight organic model compounds with various structural and functional characteristics by size exclusion chromatography combined with online OCD and ultraviolet light absorption measurement (UVD). The detection times of some low molecular weight organic compounds were much lower than expected. Elution behaviour of low molecular weight compounds (below 300 g/mol) was determined by functional groups rather than by molecular weight. Retention times of low molecular weight compounds varied between 51.9 min, close to that of humic substances, and 235 min of benzaldehyde, the double value of a standard chromatogram runtime of 120 min. Comparisons of both detectors' signals reveal that elution times and detection times are not identical. The elution sequence of various model compounds provided here facilitates identification of peaks in the low molecular weight range of SEC chromatograms. humic substance, low molecular weight compound, NOM, organic carbon detection, size-exclusion chromatography This content is only available as a PDF. © IWA Publishing 2012 You do not currently have access to this content.

Using Size-Exclusion Chromatography to Evaluate Changes in the Sizes of Au and Au/Pd Core/Shell Nanoparticles Under Thermal Treatment

In this study, we used size-exclusion chromatography (SEC) to evaluate the sizes of Au and Au/Pd core/shell nanoparticles (NPs) that had been subjected to thermal treatment, with the eluted NPs monitored through diode array detection (DAD) of the surface plasmon (SP) absorption of the NPs. In the absence of an adequate stabilizer, thermal treatment resulted in longer retention times for the Au NPs and shorter retention times for the Au/Pd core/shell NPs in the SEC chromatograms. Thus, thermal treatment influenced the sizes of these Au and Au/Pd core/shell NPs, through digestive ripening and Ostwald-type growth, respectively. In addition, the trends in the SP absorption phenomena of the NPs in the eluted samples, as measured using DAD, were consistent with the trends of their size variations, as measured from their elution profiles. In the presence of 3A-amino-3A-deoxy-(2AS,3AS)-β-cyclodextrin (H2N-β-CD) as a stabilizer, the retention times and SP absorptions of the eluted Au and Au/Pd NP samples remained constant. Thus, H2N-β-CD is a good stabilizer against size variation duration the thermal treatment of Au and Au/Pd core/shell NPs. A good correlation existed between the sizes obtained using SEC and those provided by transmission electron microscopy. Therefore, this SEC strategy is an effective means of further searching for suitable stabilizers for NPs, especially those exposed to harsh reaction conditions (e.g., in catalytic reactions).